Download Laboratory Diagnostics for Plant Pathogens of Regulatory Concern

Laboratory Diagnostics
for Plant Pathogens of
Regulatory Concern
A National Laboratory
Accreditation and Certification
System
Phil Berger
National Science Program Leader - Molecular Diagnostics &
Biotechnology
USDA APHIS PPQ
Center for Plant Health Science & Technology
NPDN Linkages
APHIS-PPQ
Cooperative Ext
Industry
NPDN
APHIS PPQ
Databases
CAPS
GPDD
NAPIS
OPIS
State Dept of Ag
(NPB)
LGU
NBIS
NPDN Critical System Requirements
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Validated Diagnostics
Plant Diagnostic Information System
– Information Management (Alerts)
– Data Security, Analysis, and Flow
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Training & Scenario Exercises
Multidisciplinary Scientific Expertise
Cross-ties to Other Networks
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Linkage with APHIS VS
National Seed Health System (ASTA)
FDA (ISO 17025)
International Regulatory Agency Contacts
• EPPO, NAPPO, CFIA, DEFRA, IPPC
– Other (NBIS)
Challenges to the NPDN
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Diversity and Numbers of Crops, Pests,
Pathogens, & Production Systems
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Over 1 billion Acres of crops, forage and forest
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Biology, Epidemiology
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Response and Recovery
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Threat Awareness
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Technology…“State of the Art”
ID/Diagnostic Challenges
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Symptoms frequently not definitive
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Morphology of pathogens
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Immunology (ELISA)
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Molecular diagnostics (PCR)
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Biochemical analyses
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Culture
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Knowledge base (something new & different)
Meeting the Challenges
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Validated Diagnostics
 Laboratory Accreditation
– Certification, Proficiency Testing
– Permits
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Harnessing New Technology
 Roles & Responsibilities
– (NPDN  Regulatory Agencies)
– Offshore Pest Information Systems
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Information Technologies
Working Definitions
➢ Accreditation: The determination that a laboratory is capable of performing
competent diagnoses.
➢ Certification: The determination that a laboratory has undergone training,
method validation, and quality assurance procedures for a specific diagnostic
method.
➢ Assay: a quantitative or qualitative test of a substance (especially an ore or a
drug) to determine its components; frequently used to test for the presence or
concentration of infectious agents or antibodies etc.
➢ Proficiency: competence in something, or knowledge of it.
.
➢ Validation: Assay validation is a process of determining the suitability of a
given laboratory method for providing useful analytical data. It describes in
mathematically quantifiable terms the performance characteristics of an assay,
identifying the source of potential errors and the likelihood of their occurrence.
(NIAID)
➢ Registration: Evaluation and recognition of a quality system that defines how
to produce the outputs (products or services) of the organization. Does not
certify or guarantee the quality of products or service for compliance with
specific specifications.
Purpose of Accreditation
To accredit laboratories in the National
Plant Disease Network (NPDN), State Dept.
of Ag., and private or commercial sector, to
carry out diagnostic tests on plant
pathogens under defined standards for
facilities, equipment, personnel training,
and certified methods.
National Plant Diagnostic Laboratory
Certification Workshop
October 12-13, 2004
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Organized by CSREES, APHIS & ARS
Attendees from NPDN, NPB, ESCOP, private industry, and
USDA (CSREES, APHIS, ARS)
Presentations:
 Validation procedures, Peter Emanuel, DOD
 Accreditation procedures, Leon Thacker, AAVLD
 [Certification procedures, Barbara Martin, NAHLN]
 Certification and Accreditation, Denis McGee, NSHA
 Accreditation procedures, Cathy Burns, FDA
 Canadian Food Inspection Agency & NAPPO, John McDonald
National Plant Diagnostic Laboratory Certification
Workshop
Desired outcomes

Understanding other laboratory accreditation and
certification systems
– What worked & what didn’t - what can we do to avoid the
same mistake(s); what are the pitfalls and how can they be
addressed
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Clarification of:
– Guiding principles for the system
– Physical and technical requirements for labs
– Procedures for accreditation and certification
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Identification of resources needed to implement the system
and major milestones in implementation
How much will it cost to implement and maintain?
National Plant Diagnostic Laboratory Certification
Workshop
October 12-13, 2004

Outcomes:
– APHIS and CSREES to collaboratively organize a
Planning/Coordinating Committee (with APHIS, CSREES,
ARS, NPDN, NPB representation)
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determine how to get the accreditation process underway and
keep things moving
provide oversight and set standards
Hire ‘Accreditation Manager’ in APHIS to develop
criteria
Hire Accreditation Developer and Implementer in
CSREES to set up and implement process, based
on specifications provided by APHIS
other …
Desirable/optimal Characteristics of a System for
Laboratory Accreditation for Plant Pathogen Diagnostics
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All labs are ISO registered
– process for documentation of non-conformances
– process for continual improvement performance
Minimum competency standards would be
established
Labs would meet regulatory requirements
There would be assurance of acceptance of
laboratory data
– Interstate assurances
– International assurances
ISO/IEC 17025-1999 Accreditation - a
framework for operating procedures, QA, etc. …
 This standard requires documentation of quality policies,
procedures, and test methods in a Quality System Manual and
is geared toward laboratories conducting routine testing.
 One of the challenges in applying ISO/IEC 17025 to the
regulatory diagnostic mandate is that the standard was
designed to suit laboratories working in the physical sciences.
 The process of pest identification goes far beyond determining
one or more characteristics by the act of separating an organism
from all others, based on a complex array of recognized
taxonomic characteristics, e.g. morphological, biochemical and
DNA sequence.
Courtesy J.G. McDonald, CFIA
ISO/IEC 17025 and Plant Pest
Identification
o ISO/IEC 17025 requires that all tests either be standard methods,
that contain the following information, or be non-standard methods,
that shall be subject to agreement with the client. The laboratory
shall also have a clear specification of the client’s requirements and
use tests that have been validated and documented appropriately.
Documentation should include at least the following:
a) appropriate identification;
b) scope;
c) description of the type of item to be tested;
d) parameters or quantities to be determined;
e) apparatus and equipment, including technical performance
requirements;
f) reference standards and reference materials required;
g) environmental conditions required and any stabilization period needed;
Courtesy J.G. McDonald, CFIA
ISO/IEC 17025 and Plant Pest Identification
(con’t)
h) description of the procedure, including:
affixing of identification marks, handling transporting, storing and
preparation of items,
checks to be made before the work is started,
checks to be made that the equipment is working properly and,
where required,
 calibration and adjustment of the equipment before each use, the method of
recording and the observations and results, any safety measures to be observed;
i) criteria and/or requirements for approval/rejection;
j) data to be recorded and method of analysis and presentation;
k) the uncertainty or the procedure for estimating uncertainty.
Courtesy J.G. McDonald, CFIA
Accreditation of a Diagnostic
Laboratory
To be accredited to perform APHIS-PPQ
validated diagnostic tests, a laboratory must
have:
Appropriate facilities and infrastructure
 Adequate and sustained institutional financial
commitment
Appropriate and properly maintained
instrumentation
Personnel (preferably permanent) with appropriate
experience and training
Training Programs
knowledge vs. skill
 An effective technician understands the method (and the science
behind it) and has skill in using it. S/he can recognize when the
method is out of specification.
 requires an ‘RO’ with appropriate background
 notification/review if RO changes --> technicians need
appropriate skill and supervision
 Designed to provide practitioners with relevant background
information (e.g., basic biology of P. ramorum)
 Basic background on the science underlying the method(s) used
for assay
 Detailed knowledge of the specific method
 Troubleshooting
 Proficiency
Proficiency
 Demonstration that the laboratory can perform the
method within specification
 Continued proficiency: what works today needs to work
tomorrow, and the next day, and the next …
 Developing proficiency panels that demonstrate skill in
performing the method can be challenging, e.g.,
 Laboratory prepared samples vs. ‘real world’ samples
 Continuous verification of panel integrity
 Internal verification and auditing of panel samples
 Experimental material cannot be used
Issues to be addressed/resolved
• Who is to be accrediting body? At present not identified, but
could be a joint function of USDA-APHIS and USDA-CSREES.
• Who will provide initial and annual funding to develop and
sustain system? Administrative unit, regional centers,
diagnostic labs…
• Is rule making required? May be necessary if regulatory
action is required by USDA-APHIS on the basis of diagnosis
made in the NPDN. Would significantly increase the time
needed to implement the system.
• Where and how does ISO fit in the plan? The system, itself,
must first be put in place. To comply with an ISO Standard,
extensive resources of time and labor are required in document
preparation. Significant fees and auditing cost are also required.
What steps should be taken to begin the
planning process
• Approval of a plan by the potential Accrediting Bodies
(CSRESS and USDA-APHIS).
• Formation of a Planning Committee. This could be a group
that evolves into the Stakeholder’s Committee.
• Appoint a Planning Director with appropriate resources and
a mandate to manage development of the system within a
certain timeframe.
Identify labs to perform
Phytophthora ramorum diagnostics
Spring 2005 December ‘04!
Goal: Up to ca. 10 labs to perform PPQ Pr
diagnostics
APHIS will establish criteria to identify
candidate laboratories:
- Laboratory infrastructure (human,
physical, & fiscal commitment
- Conduct site visits
- Identify RO for each location
- Provide training
Questions?
Assay Validation
 Assay validation is a process of determining the
suitability of a given laboratory method for providing
useful analytical data. It describes in mathematically
quantifiable terms the performance characteristics of
an assay, identifying the source of potential errors
and the likelihood of their occurrence.
(NIAID)
Common misconceptions
Assay Validation  Assay Optimization 
Assay Qualification
A Validated Method is NOT necessarily a “tight” method
Repeating an assay a number of time does not constitute
validation
source: FDA
Assay Validation Parameters:
Accuracy
Precision
Limit of Detection
Method
Validation
Limit of Quantitation
Specificity
Linearity and Range
Ruggedness/Robustness
System Suitability
source: FDA
Accuracy
Accuracy is the measure of exactness of an analytical
method, or the closeness of agreement between the
measured value and the value that is accepted as a
conventional true value or an accepted reference value.
The determination of Accuracy usually requires a “gold
standard” or an accepted method to which a new method
can be compared
- no gold standard available
- no accepted method available
source: FDA
Future Diagnostics
 Real-Time PCR
 Multiplex
 Field deployable instrumentation & kits
 Microarray hybridization
 e.g., diagnostics for mollusks; pathogen arrays
 Immunological methods
 Biosensors
 volatile organic compounds
 Gene fragment analyses
 Improved diagnostics
 Biogeographic analyses
Safeguarding US Agriculture
Pink
Bollworm
eradication
Sudden
Oak
Death
Citrus Canker
detection
Pink
Hibiscus
Mealybug
Ralstonia
solanaceraum
r3b2
Saltcedar
Plum pox
Gypsy Moth
Asian longhorn
beetle
Genetically modified
Fluorescent Green
PBW Larva
Silverleaf whitefly
Asian soybean
rust
PMTV
Rearing of sterile
medflies
USDA: National Plant Diagnostic Network
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David T. Kaplan & Philip H. Berger
USDA APHIS PPQ
Kitty F. Cardwell
USDA CSREES
November 5, 2004
End Game: Protect US Ag Production, Trade,
Natural Resources and Food Supply
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Early Detection  Low Impact
System requirements:
– Technology:
• Secure Communication
• Data Management
• Validated Diagnostics
• Laboratory Certification
– High Level of Collaboration
• Clarity of Roles, Responsibilities, and Authorities
• Capacity and Capabilities
• Technical Working Groups
• Training (Develop & Deliver)
Plant Disease & Pest Pathways
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Intentional Introduction (Bioterror: Select Agents)
– Smuggling
Natural Spread (Asiatic Soybean Rust)
Trade (e.g., Rsr3b2, ALB, EAB)
Tourism and Hurricanes (Citrus canker)
Trade and Unknown (Sudden Oak Death)
Propagative Material (Plum Pox)
Safeguarding of US Agriculture
and Natural Resources
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A Systems Approach:
– NPDN: Domestic Surveillance, Detection,
Identification, Surge Capacity & Reporting System
– Offshore Pest Detection Initiatives & Surveillance
Systems
– CAPS: Systematic survey and delineation of high
consequence or pests and pathogens in the US
– Emergency Preparedness (ICS)
• Detection, Response, Recovery
NPDN Summary
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Stewardship resides with USDA CSREES
There is a high level of collaboration
Development of NPDN is on-going
Challenges to the program are significant
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System
Multidisciplinary solutions required
Diversity of target
Technology
Future
– Threat awareness
– Linkages to other networks
– Technology